Active site binding loop stabilization in the subtilisin inhibitor eglin c: structural and functional studies on specifically designed mutants in complex with subtilisin and the uncomplexed inhibitor.
Adv Exp Med Biol
; 379: 43-7, 1996.
Article
en En
| MEDLINE
| ID: mdl-8796309
As known from the x-ray crystal structure in complex with a proteinase and from NMR studies, the serine proteinase inhibitor eglin c has a wedge-like shape with a hydrophobic core and a solvent exposed active site binding loop which is stabilized by a network of non-covalent core-binding loop interactions. Previous studies implied a crucial role of the P1'-residue Asp-46 for binding loop stabilization and high inhibitory potency of eglin c towards serine proteinases such as subtilisin. In the present study, the formation of specific eglin core-binding loop interactions was modulated by replacing the wildtype Asp-46 by asparagine, glutarnate and glutamine. The x-ray crystal structures of these mutants were solved in complex with subtilisin, and the inhibitory potency towards this enzyme was determined. Our results imply a reduction of inhibitory potency with declining core-binding loop interactions. We succeeded in crystallizing free wildtype eglin c. The 1.95 angstroms x-ray crystal structure indicates that the transition from the free to the bound form of eglin is accompanied by a concerted conformational change in the binding loop, implying an induced fit to the accessible enzyme surface. Except for the binding loop domain and a few residues on the surface of eglin, the differences observed between the uncomplexed and bound form of the inhibitor are only small.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Subtilisinas
/
Ingeniería de Proteínas
/
Inhibidores de Serina Proteinasa
/
Serpinas
/
Estructura Secundaria de Proteína
Idioma:
En
Revista:
Adv Exp Med Biol
Año:
1996
Tipo del documento:
Article
País de afiliación:
Suiza
Pais de publicación:
Estados Unidos